Quercetin supplementation promotes recovery after exercise-induced muscle damage: a systematic review and meta-analysis of randomized controlled trials

Quercetin (Q) is one of the most frequently consumed flavonoids in the human diet. The purpose of this systematic review and meta-analysis was to determine the effects of Q supplementation on muscle damage, muscle soreness and biochemical markers of inflammation, antioxidant capacity and oxidative stress after intense exercise. A literature search of SPORTDiscus, PubMed, Web of Science and Scopus was performed from inception to May 31, 2022. Forest plots were generated with fixed or random-effect models and standardized mean differences (SMD). Data extraction and quality assessment were performed independently by two authors. After application of the inclusion and exclusion criteria, 13 studies with a total of 249 sedentary to well-trained participants were included. For all studies there were some concerns about the risk of bias. All but one study used a supplementation dosage of 1000 mg/day. Q supplementation accelerated recovery of muscle function and significantly decreased muscle soreness 0/24 h after exercise (SMD: -1.33; p = 0.03), creatine kinase levels 24/48 h after exercise (SMD: -1.15; p = 0.02), and post-exercise oxidative stress (SMD: -0.92; p = 0.03). However, Q supplementation had no effect on IL-6 concentration. Q supplementation with a dose of 1000 mg/day for periods of more than seven days and a maximum of 12 weeks appears to be a safe and efficacious strategy to reduce muscle damage and muscle soreness, as well as to enhance recovery after intense exercise in sedentary to well-trained young men. Systematic review registration: PROSPERO CRD42021266801.

INTRODUCTION quercetin (Title) AND (muscle damage OR oxidative OR recovery OR muscle pain OR antioxidant OR inflammation OR soreness OR performance (Title)) AND (sports OR exercise OR train* OR athletes (Title/Abstract)). The search was limited to English language and journal articles. The search strategy is depicted in Figure 1.

Inclusion and exclusion criteria
The studies included in this systematic review and meta-analysis met the following inclusion criteria: (i) research conducted with human participants, (ii) original articles in peer-reviewed publications, (iii) original studies that investigated the effect of Q supplementation intervention on muscle damage and recovery after a protocol to induce muscle damage, (iv) research conducted with one control/placebo (PLA) group, and (v) articles published in the period from inception to the end of May 2022. Exclusion criteria were: (i) research conducted with animals, (ii) non-English articles, (iii) studies that involved other interventions in addition to Q supplementation, (iv) systematic reviews or meta-analyses, and (v) studies that reported results inadequately or without adequate statistical analysis.

Data extraction
The following data were extracted from each study: first author name, year of publication, intervention and placebo group characteristics, the dosage of supplements, supplementation duration, exercise protocol to induce muscle damage, and the effects of supplementation on functional measures, muscle soreness and biochemical markers of muscle damage, inflammation, antioxidant activity and oxidative stress.

Quality assessment
The quality of the selected articles was assessed by two independent reviewers (D.R.O. and A.J.B.R.), using the Cochrane Collaboration risk of bias tool, which covers bias in six domains: selection bias, performance bias, detection bias, attrition bias, reporting bias, and other bias. Within each domain, assessments are made for one or more items, which may cover different aspects of the domain, or different outcomes [32]. Discrepant results were resolved through discussion.

Statistical analysis
Review Manager software, version 5.4.1 (Cochrane Collaboration, Oxford, UK) was used to build the forest plot graphs and carry out the statistical analysis. The standardized mean difference (SMD) with a 95% confidence interval (CI) was used to present the results because units of measurements differed across studies. In the case of articles that reported the standard error of the mean (SEM), the standard deviation (SD) was calculated using the following formula: SD = SEM × sqrt (n), where n is the number of subjects. When necessary, the data were calculated from figures via WebPlotDigitizer. It is a program that converts graphical data to numerical data through manual plotting with high reliability [33] and has been used in other meta-analyses [34,35].
actions. It is characterized by a temporary loss of force production and delayed onset muscle soreness (DOMS), and usually has a negative impact on exercise performance [16,17]. EIMD has been described as a two-phase process [17,18]. The first phase involves structural damage to myofibers caused by the mechanical work performed by the muscles, and an increase in the production of ROS, contributing to oxidative stress. The second phase is an inflammatory response resulting from leukocyte infiltration into the damaged tissues, initiating tissue repair and adaptation [19]. However, it increases the production of ROS and can prolong inflammation, oxidative stress and cellular damage, which may delay the complete recovery of muscle function [14].
There is growing interest in the consumption of anti-inflammatory and antioxidant food supplements to reduce inflammation and enhance recovery after exercise [20]. Although some evidence suggests that antioxidant supplements may hinder the specific cellular adaptations to exercise [21], a negative impact of antioxidant supplementation on exercise training adaptation has not been reported with natural antioxidant foods [22]. Specifically, supplementation with foods containing a high concentration of phenolic compounds, such as tart cherry, pomegranate, or curcumin, has been widely used in sports to enhance performance, attenuate symptoms from EIMD, and accelerate recovery [20,[23][24][25][26].
A large body of scientific literature has investigated the effects of Q supplementation on exercise performance, based on its antioxidant and anti-inflammatory properties [27,28]. However, the results are contradictory and two systematic reviews and meta-analyses on Q supplementation and endurance performance draw different conclusions [29,30]. The effects of Q supplementation on recovery after EIMD have also been extensively studied, but the results appear, likewise, inconclusive. However, to the best of our knowledge, no systematic review or meta-analysis has been performed on this subject. Therefore, this systematic review and meta-analysis was designed to determine the effects of Q supplementation on muscle damage, muscle soreness and biochemical markers of inflammation, antioxidant capacity, and oxidative stress after intense exercise based on the existing literature.

Search strategy
This systematic review and meta-analysis was designed according to the guidelines of the Preferred Reporting of Systematic Reviews and Meta-Analyses (PRISMA) statement [31]. The protocol was registered at PROSPERO, an international database of prospectively registered reviews in health and social care (CRD42021266801).

Publication bias
Potential publication bias was not evaluated because there were fewer than 10 studies included in each meta-analysis and this number is usually chosen as a threshold value to test publication bias [37].

Search results
The literature search provided a total of 129 articles identified through the combined descriptors. After examination of the titles, 33 articles were excluded because they were meta-analyses or they were not conducted in humans. After the elimination of duplicates, 37 articles were selected for abstract screening, of which 16 were excluded for not being conducted in humans, not having a PLA group, carrying out a supplementation other than solely Q, not studying recovery, or for being systematic reviews. Twenty-one studies were then selected for full-text reading, and 8 of these were excluded for not performing an exercise protocol to induce muscle damage, for reporting results inadequately, or for inadequate statistical analysis. The final number of studies included in this systematic review and meta-analysis was 13 [38][39][40][41][42][43][44][45][46][47][48][49][50]. A summary of the search process is depicted in Figure 1.

Study Functional measures and muscle soreness
Biochemical markers of muscle damage, inflammation and oxidative stress

Study characteristics
The characteristics of the included studies are summarized in Table 1.
All selected studies were randomized placebo-controlled trials, except for that of Demirci [43], who did not mention whether participants were allocated at random. Six trials had a parallel-group design and the other 7 had a cross-over design. The total number of participants was 249 with sample sizes ranging from 10 to 20 participants in each group, except for Sholten and Sergeev [42], who only had 5 participants in the Q group and 3 in the PLA group. Only 5 studies [44,[46][47][48]50] carried out an a priori statistical power analysis.
Therefore, the other studies may not have used adequate sample sizes.
All selected studies were conducted on healthy subjects, 11 were conducted on men, 1 on women, and 1 on both women and men.
The mean age of the participants ranged from 14.90 ± 0.12 to 29.1 ± 2.4 years. Nine studies evaluated the effects of Q supplementation on trained subjects, 3 on sedentary or low to moderately active subjects, and 1 did not mention the fitness level of participants. The protocol to induce muscle damage differed greatly across the included studies. One study evaluated the effects of a single dose of 1000 mg of Q taken 3 h before the protocol to induce muscle damage [46]. The remaining studies evaluated the effects of regular Q ingestion, for more than 7 days before exercise, with a dose of 1000 mg/day except for the study of Demirci [43], who used 500 mg/day.

Methodological quality
Details of the methodological quality of the studies included are shown in Figure 2. The generation of the randomization sequence and the method of concealment were not described in any of the studies. Only one study [50] had a single blind design and one study [43] did not provide information for blinding of participants
Four studies [46][47][48][49] found a significant improvement in the Q group after supplementation in any of the variables measured after exercise or throughout the entire recovery period, when compared to the PLA group ( Table 2). Four of those studies also measured muscle soreness [41,[46][47][48], but only Bazzucchi et al. [47] observed a trend of lower muscle soreness values after exercise in the Q group (Table 2).
Meta-analysis indicated that soreness significantly decreased

Effects of Q supplementation on markers of inflammation
Seven studies [38-41, 45, 49, 50] measured one or more markers of inflammation in plasma or serum. Gholami and Ardestani [45] found a significantly lower plasma concentration of tumour necrosis factor alpha (TNF-α) in the Q group after exercise, and Sgrò et al. [49] and Tsao et al. [50] observed lower plasma interleukin-6 (IL-6) levels at some point of the recovery period. The details of the effects of Q supplementation on markers of inflammation are summarized in Table 2.

FIG. 2.
Methodological quality assessment of included trials.
Forest plots presenting the impact of Q supplementation on antioxidant activity and oxidative stress, as well as the relative weight of each study, are shown in Figures 4b and 4c.

DISCUSSION
Quercetin is one of the most frequently consumed flavonoids in the human diet and is widely distributed in fruits and vegetables. It has marked anti-inflammatory properties, as well as antioxidant activity, and exerts a protective effect against lipid peroxidation [51,52].
Recent studies have investigated whether quercetin can reduce the extent of muscle damage and accelerate recovery after exercise in humans, but the results are not conclusive.
Consumption of Q is generally recognized as safe. According to Andres et al. [53], only mild adverse effects have been reported following high supplemental doses of isolated Q up to 1000 mg/day for a maximum duration of 12 weeks in adults. However, published data on the safety of long-term supplementation of more than 12 weeks with dosages of more than 1000 mg/day of isolated Q are currently not available, and long-term supplementation with a high dose of Q could have adverse effects in humans [53].
To the authors' knowledge, this is the first systematic review and meta-analysis to examine the effectiveness of Q supplementation for recovery after EIMD in humans. Eleven studies met our inclusion criteria, involving a total of 249 participants. Our findings suggest that Q supplementation may accelerate recovery of muscle function and attenuate muscle damage and muscle soreness following strenuous exercise. Our results also suggest that Q supplementation is associated with reduced oxidative stress and increased antioxidant capacity. Therefore, Q supplementation is a good strategy to accelerate the recovery of muscle function.

Effects of Q supplementation on functional measures and muscle soreness
Four of the five studies that analysed functional measures [41,[47][48][49] used supplementation for 14 days [41,47,48], whereas the remaining study used a single dose 3 hours before exercise [46]. Bazzucchi et al. [47,48]  Therefore, it appears that Q supplementation is an effective strategy for recovery of functional measures after EIMD, even with a single dose before exercise, in sedentary to moderately active young athletes.
These results may have been due to a higher neuromuscular efficiency after Q supplementation, because Q is an adenosine receptor antagonist, which improves nerve transmission and, consequently, increases muscular strength and reduces the perceived effort of exercise and fatigue [54,55]. Our findings are broadly in agreement with those obtained in other systematic reviews on nutritional supplements [20,25,56]. can be compared with meta-analyses conducted on other ergogenic aids. Rhim et al. [35] reported that supplementation with citrulline significantly reduced feelings of muscle soreness 24 h after exercise, with an ES of -0.99, and no significant association was detected 48 h after exercise between citrulline ingestion and muscle soreness. Lv et al. [57] found that supplementation with omega-3 polyunsaturated fatty acid significantly reduced muscle soreness 48 h days after eccentric exercise, with an ES of -0.93.
Finally, Jones et al. [58] reported that nitrate-rich beetroot juice significantly reduced muscle soreness 48 h after exercise, with an ES of -0.56. Therefore, Q may be more effective for reducing muscle soreness than other nutritional supplements, because of its analgesic effect, which is dependent on many mechanisms including nitric oxide production, activation of γ-aminobutyric acid and serotonin receptors [59], and inhibition of pro-nociceptive cytokine production and oxidative stress [60].

Effects of Q supplementation on markers of muscle damage
The three studies that analysed plasma LDH [47][48][49]  Our results may be explained by the role of Q in protecting muscle fibres from damage so that the extent of cell membrane disruption is lower, which attenuates the increase of CK [47]. However, the precise mechanism of this protective effect is still unclear.

Effects of Q supplementation on markers of inflammation
The meta-analysis on IL-6 levels showed no differences between the Q and PLA groups after exercise. Our results are in line with the meta-analysis of nitrate-rich beetroot juice supplementation conducted by Jones et al. [58], who reported no significant differences in IL-6, IL-8, or TNF-a between the experimental and placebo groups after exercise. Likewise, even though Hill et al. [62] observed that tart cherry supplementation significantly decreased IL-6 after exercise, the overall ES was small. Quercetin promotes recovery after exercise extent, increasing antioxidant capacity. This is potentially due to the fact that Q is the most potent scavenger of ROS within the flavonoid family [64] because it has an optimal chemical structure for free radical scavenging, particularly because of the presence and location of the hydroxyl substitutions and the catechol group in the Bring [65,66]. In addition, Q increases the body's antioxidant capacity by regulating the levels of glutathione, which is necessary for the enzyme SOD to catalyse the decomposition of hydrogen peroxide to non-toxic H 2 O [67]. Despite Q supplementation acting directly in the secondary damage process via its antioxidant potential, it is possible to speculate that Q supplementation before exercise can reduce ROS production and limit its potentially harmful effect on cells, increasing the cell membrane resistance to the same mechanical stress [68] and therefore reducing the amount of damage caused by exercise.

Other considerations and practical applications
The results of the study by Sgrò et al. [49] suggest a new possible reason for Q to modulate damage and recovery. It is known that insulin-like growth factors 1 (IGF-I) and 2 (IGF-II) are produced during the recovery period after EIMD and both factors play an important role in skeletal muscle regeneration and remodelling [69,70]. The authors analysed the influence of Q supplementation on circulating levels of anabolic IGF-I and IGF-2, and they concluded that Q supplementation is a good strategy to promote recovery after EIMD because it increases plasma levels of anabolic factors IGF-I and IGF-II [49].
New studies are needed to support their conclusion.
Some practical applications can be drawn from this study: 1) Regular quercetin supplementation is a good strategy to reduce muscle damage and muscle soreness caused by strenuous exercise. 2) Regular quercetin supplementation also leads to a reduction of oxidative stress and increases, to some extent, antioxidant capacity. 3) One week of quercetin supplementation in doses of 1000 mg/day seems to be enough to obtain positive results.

Study limitations
There are several limitations to the current study. First, many studies did not report the necessary data for the meta-analyses. Second, the exercise protocol varied substantially between studies, inducing different levels of muscle damage. Moreover, the different training statuses of the participants and the size of the muscles involved affected the magnitude of the muscle damage experienced. Third, most studies only included young male participants; hence, these findings are unlikely to be generalizable to females or older adults.
Fourth, most studies did not perform a power analysis, and the small sample sizes were probably insufficient to detect between-group differences. In addition, though the quality of the included studies was generally acceptable, the randomization, allocation concealment, and blinding of outcome assessment procedures were inadequately described. Lastly, it was necessary to use different parameters to conduct the meta-analyses regarding antioxidant activity and oxidative stress. Although this procedure is not uncommon in the existing literature, it must be kept in mind when interpreting the results.

CONCLUSIONS
The results of our systematic review and meta-analysis support the effectiveness of Q supplementation for reducing muscle damage and accelerating recovery after intense exercise in young male athletes.
However, a limited number of studies were included in the different meta-analyses and the ES were not significant for some variables except when certain studies were removed. Therefore, our findings should be interpreted cautiously. Despite these limitations, based on the studies included in this investigation and the potential risks discussed by Andres et al. [53], Q supplementation with a dose of 1000 mg/day for periods of more than seven days and a maximum of 12 weeks appears to be a safe and efficacious strategy to reduce muscle damage and muscle soreness, as well as to enhance recovery in sedentary to well-trained young men. Athletes may also benefit from ingesting a single dose before exercise, but only one study investigated acute supplementation, and further research is needed.
More evidence is also required to confirm the efficacy of Q supplementation for females and older adults.

Aknowledgements
The authors report no financial support for this study.